ORIGINAL ARTICLE

The end of a long controversy: systematics of the genusLimenandra (Mollusca: Nudibranchia: Aeolidiidae)

Leila Carmona • Marta Pola • Terrence M. Gosliner •

Juan Lucas Cervera

Received: 2 June 2013 / Revised: 21 July 2013 / Accepted: 22 July 2013 / Published online: 7 August 2013

� Springer-Verlag Berlin Heidelberg and AWI 2013

Abstract Limenandra Haefelfinger and Stamm 1958 is a

small genus within the Aeolidiidae with, until this paper,

only two species: Limenandra nodosa Haefelfinger and

Stamm 1958 and Limenandra fusiformis Baba 1949.

Although most recent authors have regarded Limenandra

as a junior synonym of Baeolidia Bergh 1888, recent

molecular studies have demonstrated its monophyletic

status and have rejected the circumtropical distribution

attributed to the type species, L. nodosa. The present paper

reviews the previously known species of Limenandra with

additional morphological data and describes three new

species: Limenandra barnosii sp. nov. and Limenandra

rosanae sp. nov. from the Indo-Pacific are easily distin-

guished from all other Limenandra species by their vivid

and bright colour patterns, while Limenandra confusa sp.

nov., also from the Indo-Pacific, is very similar to the

Atlantic and Mediterranean L. nodosa. The five species

differ in colouration, the size and ornamentation of the

cerata, the rhinophorial papillae, details of the reproductive

system and the number of salivary glands. Additionally,

Limenandra can be easily distinguished from other Aeoli-

diidae based on differences in the radular and receptaculum

seminis morphology.

Keywords Species complex � Cladobranchia �Molluscan diversity � Morphology � New species �Taxonomy

Introduction

Haefelfinger and Stamm (1958) erected the aeolidiid genus

Limenandra, with L. nodosa as the type species, and

transferred Baeolidia fusiformis Baba, 1949 to this new

genus. They diagnosed the genus Limenandra as elongate

animals, with papillate rhinophores, fusiform cerata

arranged in rows and monoseriate radula. Nevertheless,

many authors have considered Limenandra as a junior

synonym of Baeolidia. Odhner (in Franc 1968), for

instance, included Baeolidia, but not Limenandra in his

‘‘Spurillidae’’. Gosliner (1980) also regarded Limenandra

as a junior synonym of Baeolidia and maintained this

classification with the aeolidiids from South Africa (Gos-

liner 1985). Many recent comprehensive field guides also

report L. nodosa and L. fusiformis as Baeolidia species

(Redfern 2001; Camacho-Garcıa et al. 2005; Valdes et al.

2006; Gosliner et al. 2008). In fact, since the original

description of L. nodosa, no other species has been

attributed to this genus. Recently, the first comprehensive

phylogenetic study of the Aeolidiidae (Carmona et al.

2013) demonstrated the validity of Limenandra and its

This is CEI�MAR journal publication 30.

Communicated by H.-D. Franke.

L. Carmona (&) � J. L. Cervera

Departamento de Biologıa, Facultad de Ciencias del Mar y

Ambientales, Campus de Excelencia Internacional del Mar

(CEI�MAR), Universidad de Cadiz, Polıgono Rıo San Pedro, s/n,

Ap.40, 11510 Puerto Real, Cadiz, Spain

e-mail: leila.carmona@uca.es

M. Pola

Laboratorio de Biologıa Marina, Departamento de Biologıa,

Edificio de Biologıa, Campus de Excelencia Internacional

UAM?CSIC, Universidad Autonoma de Madrid, C/Darwin,

2, 28049 Madrid, Spain

T. M. Gosliner

Department of Invertebrate Zoology, California Academy of

Sciences, 55 Music Concourse Drive, Golden Gate Park, San

Francisco, CA 94118, USA

123

Helgol Mar Res (2014) 68:37–48

DOI 10.1007/s10152-013-0367-y

monophyletic status using molecular data. The present

study provides an in-depth look at the taxonomy of the

genus and describes three new species from the eastern

Pacific and Indo-Pacific.

Materials and methods

Literature review

A comprehensive review of the literature was conducted to

determine the valid names for the species recognized in the

molecular and morphological analyses. After the descrip-

tion of the type species, all available names for Limenan-

dra species are discussed according to the year of

publication. In the synonymy lists, references to the ori-

ginal description of the valid name and all synomyms

(basionyms and primary synonyms) and the first proposed

change of binomen (secondary synonyms) are included, but

subsequent references are not.

Source of specimens and morphology

Samples were collected by SCUBA diving with standard

sampling techniques for opisthobranchs and through the

study of museum collections. When possible, two or more

specimens of each species were examined anatomically.

Specimens were dissected by dorsal incision. Their internal

features were examined and drawn using a dissecting

microscope with the aid of a camera lucida. Special

attention was paid to the morphology of the reproductive

system and the oral and salivary glands. The buccal mass

was removed and dissolved in 10 % sodium hydroxide

until the radula was isolated from the surrounding tissue.

The radula was then rinsed in water, dried, and mounted for

examination by scanning electron microscopy (SEM).

The specimens examined are deposited in the California

Academy of Sciences, CASIZ (San Francisco, USA),

Museo Nacional de Ciencias Naturales, MNCN (Madrid,

Spain), Museo de Zoologıa de la Universidad de Costa

Rica, MZUCR (San Jose, Costa Rica) and Zoologische

Staatssammlung Munchen, ZSM (Munich, Germany).

Results

Systematics

Nudibranchia Cuvier 1817

Cladobranchia Willan and Morton 1984

Family Aeolidiidae Gray 1827

Genus Limenandra Haefelfinger and Stamm, 1958. Type

species: L. nodosa Haefelfinger and Stamm, 1958.

Diagnosis of the genus Limenandra according to Hae-

felfinger and Stamm (1958).

Body thin and elongate; foot corners tentaculiform;

rhinophores papillate; cerata arranged in 8–25 rows,

fusiform and rounded; radular teeth pectinate without a

central cusp; masticatory edge of jaw smooth; ptyaline

glands (here considered as oral glands) elongate.

Limenandra nodosa Haefelfinger and Stamm 1958

(Figs. 1a, 2a, 3a, b, 4a)

Limenandra nodosa Haefelfinger and Stamm 1958: 418,

Fig. 1

Baeolidia nodosa (Haefelfinger and Stamm 1958):

Gosliner 1980: 66, Fig. 19

Material examined MNCN 15.05/63448, one specimen,

dissected, immature, 13 mm in length alive, France, Cap

Ferret, 21.vii.10, collected by Marina Poddubestkaia; CA-

SIZ 186792, one specimen, dissected, immature, 2 mm in

length preserved, Portugal, Funchal District, Madeira,

collected by Peter Wirtz 18.ix.11; MZUCR 8344, one

specimen, dissected, 4 mm in length alive, Costa Rica,

Gandoca-Manzanillo, Provincia de Limon, Punta Mona,

collected by Yolanda Camacho-Garcıa, 16.v.11; CASIZ

184521, one specimen, 8 mm in length alive, Bahamas,

Abaco, collected by Colin Redfern, 02.viii.00; MNCN

15.05/63449, one specimen, dissected, 8 mm in length

alive, Bahamas, Abaco, collected by Colin Redfern,

09.viii.00; MNCN 15.05/63450, one specimen, immature,

4.5 mm in length alive, Bahamas, Abaco, collected by

Colin Redfern, 18.viii.03.

Geographical distribution Originally described from the

Mediterranean (Haefelfinger and Stamm 1958), this species

is also known from the Atlantic coast of France (present

study), the Madeira Is. (present study), the Canary Is.

(Cervera et al. 2004), Bahamas (Redfern 2001), the

Caribbean (Valdes et al. 2006), the Atlantic coast of Costa

Rica (present study) and Curacao (Er. Marcus and Ev.

Marcus 1970).

Those specimens from the Pacific reported as L. nodosa

(Gosliner 1980; Camacho-Garcıa et al. 2005; Gosliner

et al. 2008) are here identified as L. confusa sp. nov., a

cryptic species of L. nodosa (see below).

External morphology (Figs 1a, 2, 3a, 4): The body is

elongate and slender, tapering close to the tail. The anterior

margin of the foot is rounded. The tentaculiform foot

corners have a deep groove across width (Fig. 2). The body

is dull olive green with small white spots over the entire

surface. Some specimens are whitish background with dull

olive green spots scattered over the notum. The dorsal

surface of the body from head to tail has series of con-

centric circles that are, from outside to inside, white,

yellow (sometimes absent), red and white in the centre. The

38 Helgol Mar Res (2014) 68:37–48

123

number of circles varies with the size of the animal. The

rhinophores are dull green with numerous small white

spots. The posterior side of the rhinophores has sparse,

elongate and occasionally bifurcated papillae (Fig. 3a).

The apex is rounded, white or cream. The oral tentacles are

elongate and longer than the rhinophores. The oral tenta-

cles and the foot corners have the same colour as the body

with opaque white spots.

The cerata are dorsoventrally flattened and some of them

are papillate (Fig. 4). They have the same colour as the

body covered by white spots. Some specimens have light

green pigment over the ceratal surface and one pink spot on

the anterior side, close to the base. The cerata are arranged

in up to eight rows, from just behind the rhinophores to the

end of the foot. The first two rows are very close together.

The cerata of rows IV, VI, VIII and X (sometimes absent)

Fig. 1 Photographs of the living animals. a L. nodosa, specimen

from France, Cap Ferret, photograph by Marina Poddubetskaia,

MNCN 15.05/63448; b L. fusiformis, specimen from Japan, photo-

graph by Shin Minowa, CASIZ 184526; c L. barnosii sp. nov.,

specimen from the Philippines, photograph by Terrence M. Gosliner,

CASIZ 177682; d L. confusa sp. nov., specimen from the Philippines,

photograph by Terrence M. Gosliner, CASIZ 181280

Helgol Mar Res (2014) 68:37–48 39

123

are much longer than the rest of the cerata. Each row

contains 1–5 cerata, decreasing in size towards the foot.

The anus is cleioproctic and is located behind the third

right row. The gonopore is located between rows II and III

on the right side.

Anatomy The jaws have a denticulate masticatory border

(Fig. 5a). The radular formulae are 5 9 0.1.0 (CASIZ

186792), 6 9 0.1.0 (MZUCR 8344) and 7 9 0.1.0 (MNCN

15.05/63449). The radular teeth have a total of 21–27

moderately broad denticles (MNCN 15.05/63449)

(Fig. 5b). The teeth are progressively smaller to the pos-

terior region of the radula. The oral glands lie dorsolater-

ally to the buccal bulb. They are elongate, tapering towards

the proximal end.

The reproductive system is diaulic (Fig. 6a). The pre-

ampullary duct widens into the conspicuous but short

ampulla. The postampullary duct narrows before dividing

into the oviduct and the vas deferens. The long vas def-

erens enters the wider proximal portion of the penial sac,

which contains the unarmed penial papilla. The short

oviduct gives off a duct to the circular receptaculum se-

minis before entering the female gland. The proximal part

of the duct to the receptaculum appears to have a swelling

of unknown function. The vagina opens ventral to the

penis.

Remarks Haefelfinger and Stamm (1958) erected this

new species based on 60 specimens found at Villefranche

and Beaulieu (France, Mediterranean). They illustrated the

external appearance of L. nodosa, its rhinophores, radular

teeth and jaws. These authors described a non-denticulate

masticatory border, but they also reported light stretch

grooves. In the specimens that we dissected, we found

smooth, denticulate (Fig. 3a) as well as striated masticatory

borders, so there is considerable intraspecific variation,

which is very common within the Aeolidiidae (Garcıa-

Gomez and Cervera 1985), but has not been reported pre-

viously for L. nodosa. The oral glands reported by Hae-

felfinger and Stamm (1958) were only observed in one of

the specimens examined in this study. Although these

differences could be produced by subjective misinterpre-

tation, similar variability has also been found in species of

Baeolidia (personal observation).

Schmekel (1970) described a penial gland in the

reproductive system of L. nodosa. This species together

with Baeolidia moebii (identified as Berghia major) is

thought to be the only aeolidiids with this gland (Gosliner

1985). However, we could find no trace of a penial gland in

any of the specimens examined here (Fig. 4a). Schmekel

(1970) considered that the second and small lobe of the

Fig. 4 Detailed view of the papillate cerata of L. nodosa and L.

confusa. Scale bar 2 mm

Fig. 3 Different papillate rhinophores in Limenandra. a Scale bar

3.5 mm; b from Haefelfinger and Stamm (1958)

Fig. 2 Ventral view of the anterior margin of the foot. Scale bar

0.5 mm

40 Helgol Mar Res (2014) 68:37–48

123

receptaculum seminis (Fig. 6a) was a wider part of the duct

of the receptaculum seminis. This feature, which was

described as a ‘‘loop’’ by Schmekel and Portmann (1982),

is corroborated by our study.

Our L. nodosa specimens clearly match the available

descriptions (Haefelfinger and Stamm 1958; Templado

1982; Schmekel and Portmann 1982). Only size varies.

While those specimens collected by Haefelfinger and

Stamm (1958) reached 25 mm, our specimens are 13 mm

maximum. Moreover, so far, it is the only Limenandra

species reported from the Mediterranean and Atlantic.

This restricted distribution distinguishes L. nodosa from

its sibling species from the Pacific (see L. confusa

remarks).

Fig. 5 Scanning electron photographs. a, b L. nodosa; a detailed

view of the masticatory border (CASIZ 186792), scale bar = 2 lm;

b radular teeth (MNCN 15.05/63449), scale bar 50 lm; c, d L.

fusiformis (CASIZ 184526); c detailed view of the masticatory

border, scale bar 10 lm; d radular teeth, scale bar 10 lm; e, f L.

barnosii sp. nov. (CASIZ 177682); e detailed view of the masticatory

border, scale bar 2 lm; f radular teeth, scale bar 20 lm

Helgol Mar Res (2014) 68:37–48 41

123

Limenandra fusiformis (Baba 1949) (Figs. 1b, 2, 3b, 5c,

d, 6b)

Baeolidia fusiformis Baba 1949: 184, pl. L, Fig. 169

Limenandra fusiformis (Baba 1949): Haefelfinger and

Stamm 1958, 421

Material examined CASIZ 184526, two specimens, dis-

sected, 20 mm in length alive, Japan, Chiba Prefecture,

Myogane-misaki Cap, collected by Shin Minowa,

29.vii.10; CASIZ 190512, one specimen, dissected, 3 mm

in length preserved, United Arab Emirates, East Coast, -

Gulf of Oman, collected by Carole Harris February 2005.

Geographical distribution Originally described from

Japan (Baba 1949; Nakano 2004; Ono 2004), it is also

known from the Red Sea (Yonow 2000, 2008).

External morphology (Figs. 1b, 2, 3b): The body is thin

and slender, tapering close to the tail. The anterior margin of

the foot is rounded. The foot corners are tentaculiform and

have a deep groove across width (Fig. 2). The body colour is

pale brown with bright yellow spots over the dorsum. The

dorsal surface of the body from head to tail has series of

concentric circles that are, from outside to inside, bright

yellow, pale brown and white in the centre. The number of

circles depends on the size of the animal. The rhinophores

are ashy brown with small bright yellow spots. They are

densely papillate on posterior side and the white apex is

rounded. These papillae are elongate, not uniform in size,

and occasionally bifurcated (Fig. 3b). The oral tentacles are

elongate and longer than rhinophores. Both oral tentacles

and foot corners are brownish with opaque yellow patches.

The cerata are moderately long, fusiform and rounded.

Some cerata are noticeably longer than the rest. Their

tips are sharply recurved inwards. The cerata have the

same colour as the body colour covered by bright yellow

spots, with white cnidosacs. They are arranged in up to

fifteen rows, from just behind the rhinophores to the end

of the foot, separated by a space. Each row contains

5–11 cerata, decreasing in size towards the foot. The

cleioproctic anus is located behind the third right row.

The genital opening is placed between rows II and III on

the right.

Anatomy The masticatory edge of the jaws is devoid of

denticles (Fig. 5c). The radular formula is 8 9 0.1.0

(CASIZ 184526, 20 mm). The radular teeth have a total of

15–56 moderately broad denticles (Fig. 5d). The teeth are

Fig. 6 Reproductive system.

a L. nodosa, specimen from

Bahamas (MNCN 15.05/

63449), scale bar 0.5 mm; b L.

fusiformis, specimen from

United Arab Emirates (CASIZ

190512), scale bar 0.5 mm; c L.

barnosii sp. nov., specimen

from the Philippines, (CASIZ

177682), scale bar 0.5 mm; d L.

confusa sp. nov., specimen from

Midway Island (CASIZ

088358), scale bar 0.75 mm;

e L. rosanae sp. nov., specimen

from the Marshall Islands

(CASIZ 184522), scale bar

1 mm. am ampulla, fgm female

gland mass, ps penial sac, rs

receptaculum seminis, va

vagina, vd vas deferens

42 Helgol Mar Res (2014) 68:37–48

123

progressively smaller to the posterior region of the radula.

The oral and salivary glands are absent.

The reproductive system is diaulic (Fig. 6b). The pre-

ampullary duct widens into a very long and convoluted

ampulla. The postampullary duct forks into the oviduct and

vas deferens. The short and wide vas deferens enters the

wider proximal portion of the penial sac, which contains

the unarmed penial papilla. A long oviduct connects to a

large and ball-shaped receptaculum seminis with a basal

swelling. The remaining portion of the oviduct departs

from the base of the receptaculum and enters the female

gland. The vagina opens ventral to the penis.

Remarks Limenandra fusiformis was first described under

the name of B. fusiformis (Baba 1949). Baba (1949) erected

this species based on two Japanese specimens. Although he

described the colour of the living animal and the buccal

bulb, he did not illustrate the radular teeth or the repro-

ductive system. When Haefelfinger and Stamm (1958)

erected the genus Limenandra, they transferred B. fusifor-

mis to this new genus. However, since the validity of

Limenandra has been questioned for many years (e.g.

Gosliner 1980, 1985), this species has been usually

reported as B. fusiformis instead of L. fusiformis.

The anatomy of our specimens agrees well with the

original description by Baba (1949). The long cerata have

in L. fusiformis could be considered as diagnostic for this

species since they do not occur in the other species

described here. Additionally, the elongate ampulla of the

reproductive system, which is described for the first time

here, also distinguishes this species from the remaining

species of the genus. Specimens from the Red Sea have

some morphological differences from the Japanese speci-

mens, such as cream body colour, so clearly, further

material is required in order to confirm the presence of

L. fusiformis in the Red Sea.

Limenandra barnosii sp. nov (Figs. 1c, 2, 3b, 5e, f, 6c)

Baeolidia sp. 3: Gosliner et al. (2008), 406

Limenandra sp. C: Carmona et al. (2013), 6

Type material Holotype: CASIZ 177682, one specimen,

dissected, 30 mm in length preserved, Philippines, Luzon

Island, Batangas Province, Balayan Bay, Matotonngil Point,

collected by Terrence M. Gosliner, 19.iv.08. Paratype:

CASIZ 182761, one specimen, dissected, 40 mm in length

preserved, Philippines, Luzon Island, Batangas Province,

Balayan Bay, Anilao, collected by Roger Steene, 16.v.10.

Type locality and habitat Balayan Bay, Batangas Prov-

ince, Luzon Island, Philippines. Found in shallow water, in

areas of mixed sand and rubble, feeding on the anemone

Alicia sansibarensis.

Geographical distribution So far, only known from the

Philippines, Indonesia and New Caledonia (Gosliner et al.

2008; present study).

Etymology This species is dedicated to Mohamed Bar-

nosi, grandfather of the first author of this paper.

External morphology (Figs. 1c, 2, 3b): The body is

elongate and slender, tapering close to the tail. The anterior

margin of the foot is rounded. The tentaculiform foot

corners have a deep groove across width (Fig. 2). The body

colour is pale pink with lime green pigment all over the

dorsum. The dorsal surface of the body from head to tail

has series of concentric circles that are bright lemon yellow

and pinkish purple inside. The number of circles varies

with the size of the animal but up to fifteen. The edge of the

foot alternates transverse translucent and lime green bands,

which go from head to tail. The bases of the pink rhino-

phores are very close. They are densely papillate on pos-

terior side, presenting orange colouration on the papillae.

These papillae are elongate, not uniform in size and

occasionally bifurcated (Fig. 3b). The oral tentacles are

pinkish, elongate and longer than the rhinophores. The foot

corners have same colour as the body and two lime green

rings, the first one just below the pink tip and the second

one at the midpoint.

The cerata are moderately long, fusiform and rounded.

Their tips are slightly recurved inwards. The cerata are dark

olive green with lime green pigments scattered all over their

surface. Their distal part is pink and they have translucent

tips. The cerata are arranged in up to twenty-five rows, from

just behind the rhinophores to the end of the foot, and they

are very close together every two rows. Some rows of cerata

join over the dorsum and form an arch side-to-side with the

opposite row. Each row contains 3–15 cerata, decreasing in

size towards the foot. The anus is cleioproctic and is placed

behind the third right row. The gonopore is located between

right II and III rows on the right side.

Anatomy The buccal bulb has two jaws with a mastica-

tory border finely denticulate (Fig. 5e). The radular for-

mulae are 12 9 0.1.0 (CASIZ 177682) and 11 9 0.1.0

(CASIZ 182761). The radular teeth are wide with up to 66

moderately broad denticles (CASIZ 177682) (Fig. 5f). The

teeth are progressively smaller to the posterior region of the

radula. The oral glands lie dorsolaterally to the buccal bulb.

They are moderately long and widest in the proximal end.

Two pairs of salivary glands are present.

The reproductive system is diaulic (Fig. 6c). The pre-

ampullary duct widens into the conspicuous and elongate

ampulla. The postampullary forks into the oviduct and vas

deferens. The moderately long vas deferens enters the

wider proximal portion of the penial sac, which contains

Helgol Mar Res (2014) 68:37–48 43

123

the unarmed penial papilla. The long oviduct connects to a

ball-shaped receptaculum seminis with a basal swelling.

The remaining portion of the oviduct departs from the base

of the receptaculum and enters the female gland. The

vagina opens ventral to the penis.

Remarks Limenandra barnosii is easily distinguishable

from other Limenandra species by its bright colours. It also

differs from L. nodosa, L. confusa and L. fusiformis in the

ornamentation, colouration and size of the cerata, the num-

ber of salivary glands and the reproductive system. While L.

barnosii has smooth cerata, those of L. nodosa and L. con-

fusa are often papillate. L. fusiformis also has smooth cerata,

but some of them are significantly longer than the cerata of

L. barnosii. Moreover, only L. barnosii has two pairs of

salivary glands and a long oviduct leading from its junction

with the vas deferens to the receptaculum seminis.

Limenandra confusa sp. nov (Figs. 1d, 2, 3a, 4, 6d, 7a, b)

Baeolidia nodosa (Haefelfinger and Stamm 1958), non

Haefelfinger and Stamm 1958: Gosliner 1980, 66–69,

Fig. 19

Limenandra sp. A: Carmona et al. (2013), 6

Type material Holotype: CASIZ 088358 one specimen,

dissected, 8 mm in length preserved, Midway Islands,

South side of Sand Island, collected by Terrence M. Gos-

liner, 01.vi.93. Paratype: CASIZ 185991, one specimen,

dissected, 3 mm in length preserved, Philippines, Luzon

Island, Batangas Province, Balayan Bay, Anilao, Anilao

Harbor, collected by Alicia Hermosillo, 30.v.11.

Other material CASIZ 181280, one specimen, 3 mm in

length preserved, Philippines, Luzon, Batangas Province,

Calumpan Peninsula, collected by Peri Paleracio,

21.v.09.

Type locality and habitat South side of Sand Island,

Midway Islands. Found in shallow waters (3–10 m) under

coral rumble, feeding upon small sea anemones.

Geographical distribution This species has been reported

from the Indo-Pacific (Philippines) as well as the eastern

Pacific: the Midway Islands (present study), Hawaii (Kay

1979; Gosliner 1980; Pittman and Fiene 2012a), Mexico

(Gosliner et al. 2008), Gulf of California (Bertsch 1972)

and Costa Rica (Camacho-Garcıa et al. 2005).

Fig. 7 Scanning electron photographs. a, b L. confusa sp. nov.

(CASIZ 088358); a detailed view of the masticatory border, scale bar

2 lm; b radular teeth, scale bar 10 lm; c, d L. rosanae sp. nov.

(CASIZ 106582); c detailed view of the masticatory border, scale bar

100 lm; d radular teeth, scale bar 10 lm

44 Helgol Mar Res (2014) 68:37–48

123

Etymology The specific name refers to the mistakes pre-

viously made in identifying this species due to it close

external morphological resemblance to L. nodosa.

External morphology (Figs. 1d, 2, 3a, 4): The body is

slender and elongate, tapering close to the tail. The

anterior margin of the foot is rounded. The foot corners

are tentaculiform and have a deep groove across width

(Fig. 2). The body colour is dull olive green with small

white and cream spots over the notum. Series of con-

centric circles that are, from outside to inside, yellow, red

and white in the centre, over the notum are found. The

number of circles varies with the size of the animal. The

rhinophores are dull green with small white and/or cream

spots. The posterior side of the rhinophores has sparse

papillae (Fig. 3a). These papillae are elongate and may be

bifurcated. The apex is white or cream and rounded. The

oral tentacles are elongate, longer than rhinophores. The

oral tentacles and the foot corners are dull olive green

with opaque white spots. The cerata are dorsoventrally

flattened, some of them papillate (Fig. 4). They have the

same body colour covered by white spots. The cerata are

arranged in up to eleven rows, from just behind the rhi-

nophores to the end of the foot, the first two rows very

close together. The cerata of rows IV, VI, VIII and X

(sometimes absent) are much longer than the rest. Each

row contains 1–6 cerata, decreasing in size towards the

foot. The cleioproctic anus is placed behind the third right

row. The genital opening is located between rows II and

III on the right.

Anatomy The jaws have a smooth masticatory edge

(Fig. 7a). The radular formula is 9 9 0.1.0 (CASIZ

088358). The radular teeth have a total of 20–40 moder-

ately broad denticles (CASIZ 088358) (Fig. 7b). The teeth

are progressively smaller to the posterior region of the

radula. Oral and salivary glands are present.

The reproductive system is diaulic (Fig. 6d). The pre-

ampullary duct widens into the conspicuous and short

ampulla. The ampulla narrows again before dividing into

the oviduct and vas deferens. The moderately long vas

deferens enters the wider proximal portion of the penial

sac, where the unarmed penial papilla is housed. The short

oviduct connects to a rounded and large receptaculum se-

minis with a basal swelling. The remaining portion of the

oviduct departs from the base of the receptaculum and

enters the female gland. The vagina opens ventral to the

penis.

Remarks Apart from the salivary glands and size of the

receptaculum seminis, no morphological differences are

found between L. nodosa and L. confusa. Since the oral

glands can be easily misinterpreted for a piece of tissue or

even missed or ignored, these glands usually do not appear

in previous species descriptions. More studies are needed

in order to confirm the validity of the salivary glands as a

taxonomical character to distinguish species, but our

observations may indicate the value of this anatomical

feature.

Regarding the receptaculum seminis, L. confusa has a

significantly larger receptaculum seminis than L. nodosa.

When Gosliner (1980) reported L. confusa in Hawaii

(under the name of L. nodosa), he also illustrated a large

receptaculum seminis. Although more specimens of

L. confusa would help to confirm the consistency of this

morphological character, we consider the receptaculum

seminis as the main morphological difference between

L. nodosa and L. confusa.

Bertsch (1972) compared the characteristics of the rad-

ula, jaws, external colouration, cerata, size and rhinophores

between one specimen of L. fusiformis and two specimens

of ‘‘L. nodosa’’, one from the Mediterranean and the other

from the Gulf of California (see Table 1 of that contribu-

tion). This author detected an overlap of these morpho-

logical traits but found some variability in colouration and

shape of rhinophores. Nevertheless, Bertsch (1972) could

not determine whether ‘‘these variations are all character-

istic of one, two or three species’’. All the differences

pointed out by Bertsch (1972) have been reviewed and

rejected as no external differences were found between

L. nodosa and L. confusa. However, Carmona et al. (2013)

found genetic differences between these two species

(minimum uncorrected p-distance = 7.3 % for COI

between L. nodosa and L. confusa, see Table 2 of that

contribution). Recent studies supported by molecular

phylogenies are starting to separate taxa with ecological

or geographical differences (Claremont et al. 2011;

Johnson and Gosliner 2012). Clearly, both species have

distinct geographical patterns (Atlantic–Mediterranean for

L. nodosa and Pacific for L. confusa). Thus, in view of the

genetic, anatomical and biogeographic differences, we

consider L. nodosa and L. confusa to be two sibling cryptic

species.

Limenandra rosanae sp. nov (Figs. 2, 3b, 6e, 7c, d,

8a, b)

Limenandra sp. 1: Ono (1999a)

Limenandra nodosa (Haefelfinger and Stamm 1958),

non Haefelfinger and Stamm 1958: Ono 1999b: 168, first

photo from the top

Baeolidia sp. 2: Gosliner et al. (2008), 406

Limenandra sp.: Yonow (2008), 253

Baeolidia sp. #2: Pittman and Fiene (2012b)

Limenandra sp. B: Carmona et al. (2013), 6

Material examined Holotype: CASIZ 184522, one spec-

imen, dissected, 15 mm in length preserved, Marshall

Helgol Mar Res (2014) 68:37–48 45

123

Islands, Kwajalein Atoll, collected by Scott Johnson,

25.vi.09; Paratype: CASIZ 106582, one specimen, dis-

sected, 2.5 mm in length preserved, Philippines, Luzon,

Batangas Province, Maricaban Island, Cemetery Beach,

collected by Terrence M. Gosliner, 15.iv.96.

Other material CASIZ 184528, one specimen, dissected,

9 mm in length preserved, Marshall Islands, Kwajalein

Atoll, collected by Scott Johnson, 30.viii.09; ZSM Mol-

20052082, one specimen, dissected, 7 mm in length pre-

served, Samoa, 17.viii.05.

Geographical distribution Known from the Marshall Is.

(present study), Hawaii (Gosliner et al. 2008; Pittman and

Fiene 2012b), Samoa (present study), the Philippines

(Gosliner et al. 2008) and the Kerama Is. (Ono 1999a, b).

Also reported from the Red Sea (Yonow 2000, 2008).

Type locality and habitat Kwajalein Atoll, Marshall

Islands. Found on shallow patch reefs in the vicinity of sea

anemones.

Etymology This species is dedicated to Rosana Carvalho

Schrodl, wife of Michael Schrodl who kindly gave us the

first specimen of this species.

External morphology (Figs. 2, 3b, 8a, b): The body is

elongate and slender, tapering close to the tail. The anterior

margin of the foot is rounded. The tentaculiform foot

corners have a deep groove across width (Fig. 2). The

body colour is pearly white. The dorsal surface from head

to tail has series of concentric circles that are bright lemon

yellow (sometimes absent), bright grey and pearly white or

pearly pink inside. In some specimens, these circles

alternate with concentric circles of bright lemon yellow

Fig. 8 Photographs of the living animals. a, b, L. rosanae sp. nov. specimens from the Marshall Islands, photographs by Scott Johnson, CASIZ

106582 and CASIZ 184528, respectively

46 Helgol Mar Res (2014) 68:37–48

123

outside and bright grey in the centre. The number of circles

depends on the size of the animal but may be up to fifteen.

Small ochre or dark green/black spots together with bright

yellow patches form a continuous band on each side of the

body. The bases of the rhinophores are very close. The

rhinophores are pearly white with the small ochre or dark

green/black spots. They are densely papillate on posterior

side. These papillae are elongate, not uniform in size, and

may be bifurcated (Fig. 3b). Their apex is sharply rounded.

The oral tentacles are elongate, longer than rhinophores.

Both oral tentacles and foot corners have the same colour

as the body, presenting the small ochre or dark green/black

spots that are arranged in rings. The cerata are moderately

short, fusiform and rounded. Their tips are slightly

recurved towards the tail. The cerata are pearly white

overlaid by ochre or dark green/black spots. Some speci-

mens also have bright lemon yellow colouration on the

outer side of the cerata. They are arranged in up to eigh-

teen rows, from the rear of rhinophores to the end of the

foot, separated by a small space every two rows. Each row

contains 1–7 cerata, decreasing in size towards the foot.

The anus is cleioproctic and is located behind the third

right row. The genital aperture is placed between right II

and III rows on the right.

Anatomy The jaws have a smooth masticatory border

(Fig. 7c). The radular formulae are 4 9 0.1.0 (CASIZ

106582) and 5 9 0.1.0 (ZSM Mol-20052082). The radular

teeth have a total of 18–33 moderately broad denticles

(CASIZ 106582) (Fig. 7d). The teeth are progressively

smaller to the posterior region of the radula. The oral

glands lie dorsolaterally to the buccal bulb. They are

moderately long and widest in the proximal end. Two pairs

of salivary glands are present.

The reproductive system is diaulic (Fig. 6e). The pre-

ampullary duct widens into the wide and short ampulla,

which narrows again before dividing into the oviduct and

vas deferens. The vas deferens is usually conspicuous,

although the specimen CASIZ 106582 has a thin and

elongate vas deferens. This structure enters the wider

proximal portion of the penial sac, which contains the

unarmed penial papilla. The short oviduct connects to a

large and pyriform receptaculum seminis with a basal

swelling. The remaining portion of the oviduct departs

from the base of the receptaculum and enters the female

gland. The vagina opens ventral to the penis.

Remarks Limenandra rosanae is clearly distinguishable

from other members of the genus because of its external

colouration and its pyriform receptaculum seminis. Fur-

thermore, this species differs from L. nodosa and L. con-

fusa in having two salivary glands and in lacking long

cerata or papillae on any cerata. L. barnosii also has a pair

of salivary glands but differs in colouration.

Discussion

Phylogenetic analysis conducted by Carmona et al. (2013)

strongly supported the monophyly and validity of the genus

Limenandra, which had previously been considered a

junior synonym of Baeolidia (Gosliner 1980, 1985; Red-

fern 2001; Camacho-Garcıa et al. 2005; Valdes et al. 2006;

Gosliner et al. 2008). The original diagnosis of the genus

proposed by Haefelfinger and Stamm (1958) clearly mat-

ches with our results. Nevertheless, the description of the

rhinophoral papillae, the presence of concentric circles on

the notum, the swelling of the receptaculum seminis and

the small denticles on the masticatory border of some

specimens should to be added to the diagnosis. We there-

fore amend the following diagnosis of the genus: body thin

and elongate. Tentaculiform foot corners. Rhinophores

papillate; the papillae elongate, irregular in size and

sometimes bifurcated. Colouration pattern of concentric

circles on the notum. Cerata in 8–25 rows. They are fusi-

form, rounded and may be papillate. Radular teeth pro-

gressively smaller to the posterior region of the radula;

pectinate without a distinct central cusp. Masticatory edge

usually smooth. Receptaculum may have a basal swelling

or a swelling at the base of the duct to the receptaculum.

This study shows that this small genus Limenandra of

just five species so far has a mainly Indo-Pacific distribu-

tion, which agrees with the conclusions of Carmona et al.

(2013). Only L. nodosa does not occur in the Indo-Pacific

Fig. 9 Phylogeny of

Limenandra, based on Carmona

et al. (2013). Names on right

side of vertical bars refer to

revised classification

Helgol Mar Res (2014) 68:37–48 47

123

but is an amphiatlantic species. Figure 9 depicts the phy-

logenetic hypothesis presented by Carmona et al. (2013)

resolving the phylogenetic relationships within the genus

(names of the new species described here appear on the

right side of vertical bars). According to Fig. 9, L. rosanae

clusters in a separate basal clade. The second clade is

divided into two subclades: one with L. confusa and

L. nodosa as sister species and the second one with

L. barnosii and L. fusiformis.

Acknowledgments We are deeply grateful to all individuals who

helped to collect and provided specimens and images for this study,

including Y. Camacho-Garcıa, M. Schrodl, C. Redfern, R. Nakano,

M. Poddubetskaia, D. Behrens, C. Harris and S. Johnson. A. Sellas

helped us with some molecular laboratory work. This work was

supported by several research Grants CGL2006-05182/BOS,

CTM2008-05228-E/MAR and CGL2010-17187, Spanish Ministry of

Economy and Competitiveness (includes the early Ministry of Sci-

ences and Innovation) to J. L. Cervera; Institut Menorquı d’Estudis,

Balearic Is. to J. L. Cervera; California Academy of Sciences,

National Science Foundation (DEB 0329054 PEET) to T. M. Gos-

liner; and a Research Grant of the Malacological Society of London to

L. Carmona.

References

Baba K (1949) Opisthobranchia of Sagami Bay collected by His

Majesty The Emperor of Japan. Iwanami Shoten, Tokyo

Bertsch H (1972) Two additions to the opisthobranch fauna of the

Southern Gulf of California. Veliger 15:103–106

Camacho-Garcıa Y, Gosliner TM, Valdes A (2005) Field guide to the

sea slugs of the tropical Eastern Pacific. California Academy of

Sciences, San Francisco, CA

Carmona L, Pola M, Gosliner TM, Cervera JL (2013) A tale that

morphology fails to tell: a molecular phylogeny of Aeolidii-

dae (Aeolidida, Nudibranchia, Gastropoda). PloS One 8. doi:10.

1371/journal.pone.0063000

Cervera JL, Calado G, Gavaia C, Malaquias MAE, Templado J,

Ballesteros M, Garcıa-Gomez JC, Megina C (2004) An anno-

tated and updated checklist of the opisthobranchs (Mollusca:

Gastropoda) from Spain and Portugal (including islands and

archipelagos). Bol Inst Esp Oceanogr 20:1–122

Claremont M, Reid DG, Williams ST (2011) Evolution of corallivory

in the gastropod genus Drupella. Coral Reefs 30:977–990

Franc A (1968) Sous-classe des opisthobranches. In: Grasse PP (ed)

Traite de Zoologie. Masson, Paris, pp 608–893

Garcıa-Gomez JC, Cervera JL (1985) Revision de Spurilla neapol-

itana Delle Chiaje 1823 (Mollusca: Nudibranchiata). J Molluscan

Stud 51:138–156

Gosliner TM (1980) The systematics of the Aeolidacea (Nudibran-

chia: Mollusca) of the Hawaiian Islands, with description of two

new species. Pac Sci 33:37–77

Gosliner TM (1985) The aeolid nudibranch family Aeolidiidae

(Gastropoda, Opisthobranchia) from tropical southern Africa.

Ann S Afr Mus 95:233–267

Gosliner TM, Behrens DW, Valdes A (2008) Indo-Pacific nudi-

branchs and sea slugs. Sea Challengers/California Academy of

Sciences, Gig Harbor, San Francisco, CA

Haefelfinger HR, Stamm RA (1958) Limenandra nodosa gen. et spec.

nov. (Nudibranchia, Aeolidiidae prop.), un opisthobranche

nouveau de la Mediterranee. Vie Milieu 9:418–423

Johnson RF, Gosliner TM (2012) Traditional taxonomic groupings

mask evolutionary history: a molecular phylogeny and new

classification of the chromodorid nudibranchs. PloS One 7.

doi:10.1371/journal.pone.0033479

Kay EA (1979) Hawaiian marine shells, reef and shore fauna of

Hawaii, section 4: Mollusca. Bernice P. Bishop Museum Special

Publication 64(4)

Marcus Er, Marcus Ev (1970) Opisthobranchs from Curacao and

faunistically related regions. Stud Fauna Curacao Caribbean Isl

33:1–129

Nakano R (2004) Opisthobranchs of Japan Islands. Rutles Inc, Tokyo

Ono A (1999a) Another species of Limenandra from Kerama Islands?

[Message in] Sea Slug Forum. Australian Museum, Sydney.

Available from http://www.seaslugforum.net/find/1215. Acces-

sed November 2012

Ono A (1999b) Opisthobranchs of Kerama Islands. TBS-Britannica

Co., Ltd., Tokyo

Ono A (2004) Opisthobranchs of Ryukyu Islands. Rutles, Inc., Tokyo

Pittman C, Fiene P (2012a) Baeolidia nodosa. [Message in] Sea Slugs

of Hawaii. Available from http://seaslugsofhawaii.com/species/

Baeolidia-nodosa-a.html. Accessed November 2012

Pittman C, Fiene P (2012b) Baeolidia sp. #2. [Message in] Sea Slugs

of Hawaii. Available from http://seaslugsofhawaii.com/species/

Baeolidia-sp2-a.html. Accessed November 2012

Redfern C (2001) Bahamian Seashells: a thousand species from

Abaco, Bahamas. Bahamianseashells.com Inc, Boca Raton, FL

Schmekel L (1970) Anatomie der Genitalorgane von Nudibranchiern

(Gastropoda Euthyneura). PSZNI 38:120–217

Schmekel L, Portmann A (1982) Opisthobranchia des Mittelmeeres.

Springer, Berlin

Templado J (1982) Nuevo opistobranquio para la malacofauna

iberica. Actas II Simposio Iberico sobre los Estudios Bentos

Marino 3:225–227

Valdes A, Hamann J, Behrens DW, DuPont A (2006) Caribbean sea

slugs. A field guide to the opisthobranch mollusks from the

tropical northwestern Atlantic. Sea Challengers, Gig Harbor,

WA

Yonow N (2000) Red Sea Opisthobrachia 4: the orders Cephalaspi-

dea, Anaspidea, Notaspidea and Nudibranchia: Dendronotacea

and Aeolidacea. Fauna Arabia 18:87–131

Yonow N (2008) Sea slugs of the Red Sea. Pensoft Publ, Sofıa,

Bulgaria

48 Helgol Mar Res (2014) 68:37–48

123